The goal of our research is to understand the mechanism of translational control of protein synthesis in eukaryotic cells. We have developed a system that combines the advantages of reticulocytes to study and dissect protein synthesis and heat shock to rapidly alter translational specificity. We have found that chicken reticulocytes respond to heat shock by the increased synthesis of only one heat shock protein (HSP70) and the rapid repression of globin synthesis. The globin mRNA is neither degraded nor structurally modified in heat shocked cells and can be isolated and in vitro translated. HSP70 mRNA is transcribed in normal reticulocytes and apparently maintained in a translationally repressed state in the cytoplasm. Heat shock induces the selective synthesis of HSP70 even though there is no significant increase in the amount of HSP70 mRNA in the cytoplasm. We suggest that the increased synthesis of HSP70 is due to selective utilization of HSP70 mRNA in heat shocked cells. The selectivity may be due to heat shock induced modifications of the translational apparatus or other factors that discriminate between HSP70 mRNA and other cellular mRNAs. We expect that HSP70 mRNA contains features that are recognized by these discriminatory factors. Our system is ideally suited for studying translational control because the parameters of protein synthesis are well characterized in reticulocytes and these cells from chickens respond to heat shock by the increased synthesis of only HSP70. This provides the opportunity to focus on the expression of a single species of mRNA, that for HSP70, and the regulation of its interaction with the translational apparatus. The experiments proposed in this grant will elucidate the mechanism by which translational control operates, preferentially utilizes HSP70 mRNA and results in the selective translation of HSP70 mRNA in heat shocked cells. These goals will be accomplished by identifying the changes in the protein synthetic apparatus that affect translational specificity by using a chicken reticulocyte in vitro translation system. We will also determine whether HSP70 mRNA contains information necessary for its preferential translation by mutational analysis of the cloned chicken HSP70 gene and introduction of the modified genes into chicken lymphoid and erythroid cells. The studies will increase our knowledge about basic cellular control mechanisms, in particular how vertebrate cells respond to alterations in their environment.